At present influenza virus is one of main pathogenic microorganisms harmful to human health, which not only causes great economic loss, but also affects the society stability. The natural host of influenza virus is avian, and the avian influenza virus can break through barrier between species to infect people. So far, type A influenza virus which can infect human and cause human influenza pandemic or seasonal influenza can only be subtype H1N1, subtype H2N2 or subtype H3N2 of influenza virus. From the origin, human influenza viruses are all derived from avian influenza virus. In recent years, subtype H5 of highly pathogenic avian influenza often breaks out in poultry causing huge economic loss, and occasionally infects humans causing concentrated infections or sporadic cases in some areas. In February 2013, a new type of human-infected H7N9 avian influenza virus appeared in Anhui and Shanghai of China, which had caused 571 cases of human infection and had leaded to 212 cases of death till Feb. 23, 2015. Subtype H9N2 of influenza virus is a kind of low pathogenic influenza virus, which is common in poultry. In recent years, subtype H9N2 of influenza virus has been isolated incessantly from samples of infected patients. As early as 2004, the first case of human infection with subtype H10N7 of avian influenza virus was reported in Egypt. In Australia the cases of human infection with subtype H10N7 of virus also occurred in 2010 and 2012. In December 2013, the first case of human infection with H10N8 avian influenza virus appeared in Jiangxi province of China. In June 2013, the world's first case of human infection with subtype H6N1 of avian influenza virus was detected in Taiwan, whose viral gene sequence isolated from the patient showed that the virus was of typically low pathogenic avian influenza virus. The above studies show that there are at least eight different human-infected subtypes H1, H2, H3, H5, H6, H7, H9 and H10 of avian influenza virus.
At present, laboratory detection techniques for avian influenza virus mainly include methods like virus isolation and culture thereof, serological experiment, molecular biology experiment and immunofluorescence etc., wherein virus isolation and culture thereof are “gold standard” for the diagnosis of avian influenza virus infection. Due to various subtypes of avian influenza and many emerging variants, using the above detecting method for classifying and diagnosing avian influenza virus not only has the disadvantage of complicated operation, but also has the disadvantages of long detection period and accurate and not timely detection results easily affected by antibody etc. Although multiplex PCR has advantage of simultaneous amplification of multiple target fragments so as to realize rapid diagnosis of various kinds of pathogens and has advantage of reduced detection cost, common multiplex PCR in the amplification process are susceptible to characteristic of target gene template, primer concentration, primer proportion and interaction between PCR reagents, leading to different amplification efficiency of samples which has accuracy of detection results reduce greatly. Multiplex reverse transcription polymerase chain reaction (mRT-PCR) in GeXP multiplex gene genetic expression analysis system can solve problem of different amplification efficiency present in the common multiplex PCR by combining universal primer (labeled with fluorescent marker upstream) and specific chimeric primer (5′ end of gene specific primer is connected with universal primer sequence) to build multiplex reverse transcription polymerase chain reaction system and then using strategy of universal primer initiation. In addition, this method makes use of capillary electrophoresis to separate product, which greatly improves the sensitivity and reliability of detection result.
Currently nucleic acid detection method for detecting different subtypes of avian influenza virus is used for detecting simultaneously three different subtypes HA at most, while detection technique which can detect simultaneously eight different human-infected subtypes HA of avian influenza virus like H1, H2, H3, H5, H6, H7, H9 and H10 in one reaction has not been reported yet.
Information disclosed in the background is simply used for better comprehension of overall background of the present invention, while should not be considered as acknowledging or implying in any form that the information is general prior art for a person skilled in the art.